Cloning and characterization of flower senescence-related genes from carnation and petunia.

摘要

Senescence represents the final stage in the development of a whole plant, organ, tissue or cell and involves those processes leading to death. The flower is often the organ with the shortest life span, and as such provides a useful tissue for studying the mechanisms underlying control of senescence. Senescence of flower petals is associated with increased production of the phytohormone ethylene. This ethylene initiates and regulates the processes of programmed cell death. The onset of petal death is characterized by the expression of several senescence-related (SR) genes. Using a differential screening approach I have identified 9 unique groups of cDNA clones representing SR transcripts. These cDNA clones have been used to study specific changes in gene expression during flower senescence. In most cases the increase in abundance of SR mRNAs occurs concomitant with the ethylene climacteric. Treatment of flowers with an ethylene action inhibitor prevents petal senescence and the accumulation of SR mRNAs. Two cDNA clones, pSR132 and pSR120, representing mRNAs which accumulate in senescing carnation petals were characterized in details. The clone pSR132 was confirmed to encode a protein of approximately 35 kDa by sequence analysis and in vitro translation of hybrid-selected pSR132 mRNA. The predicted peptide sequence of pSR132 shares significant homology with enzymes involved in the formation of C-P bonds in the biosynthesis of phosphonates, CPEP and PEP mutase, from Streptomyces hygroscopius and Tetrahymena pyriformis, respectively. C-P bonds are found in a wide range of organisms, but their presence or formation in higher plants has not been investigated. The nucleotide and predicted protein sequences of pSR 120 are highly homologous to ACC oxidase which catalyzes the conversion of ACC to ethylene. A petunia ACC oxidase cDNA, pPETEFE, was isolated by screening a cDNA library prepared from mRNA of senescing petunia corolla. RNA gel blot analysis indicates the mRNA detected by pPETEFE increases in abundance in senescing corollas following pollination. In order to identify the DNA sequences and cellular factors which regulate the expression of ACC oxidase during floral senescence, I have isolated 13 genomic clones which share sequence homology with the petunia ACC oxidase cDNA. Restriction mapping indicates these represent two distinct regions of the genome. One genomic region represented by several overlapping cloned DNA fragments contains a cluster of two ACC oxidase genes. Sequence analysis indicated these two genes are very similar and likely arose by gene duplication.